Mycoplasmas are important pathogens with usually strict natural host tropism and poorly understood virulence mechanisms. The broad objective of this work is increased understanding of mycoplasmal host-range and virulence determinants through comparison of conserved and variable features of mycoplasmal genomes. Comparative genomics is a new alternative to the usual tools of molecular biology to elucidate the genetic bases of mycoplasmosis. Mycoplasma alligatoris merits priority in that effort because in some hosts it causes hyperacute lethal disease, but it is merely commensal in others. The specific aims are to assemble a draft map of the M. alligatoris genome by alignment with that of closely-related Mycoplasma pulmonis, and to identify candidate genes involved in host tropism and virulence. The working draft will be assembled by using automated high-throughput sequence assembly methods. Sequence alignments will be validated by comparative synteny and PCR-based gap closure. Coding sequences will be identified by interpolated Markov models trained with mycoplasmal sequences and cataloged according to their public database matches. Candidate host-range genes may be recognized by overall similarity to known adhesins or by signature ligand-binding motifs. Virulence factors may include homologs to cell-surface or secreted hydrolytic enzymes of other mollicutes, or to pathogenic determinants present in other bacteria. The sequences of homologs whose known biological roles or predicted properties are consistent with host cell-surface ligand-binding or the pathogenic effects of M. alligatoris will be characterized in detail. The data will be integrated in a representation of the complement of adhesion, transport, and metabolic pathways of M. alligatoris validated by the biologically well-documented metabolic map of the mollicutes. The ease of PCR-based cloning and the accessibility of bioinformatics software make this an ideal project to engage undergraduate students in innovative biomedical research.